The list of mammalian disease processes in which oxygen radicals and their cascaded reactive radicals are implicated continues to grow. In order to examine the structural identities of these species and the mechanism of the oxidative modification of biomolecules, we investigated the effects of superoxide dismutase (SOD) and transition metal ions on the formation of these radicals. Electron Paramagnetic Resonance (EPR) spectroscopy and a spin trapping method were employed for the studies. The SOD catalyzes the reaction of superoxide dismutation and thus protects biomolecules from the superoxide toxicity. However, its own reaction product hydrogen peroxide is also toxic and is known to inactivate Cu,Zn-SOD itself. We have found in this study that during the inactivation of Cu,Zn-SOD by hydrogen peroxide, "free" -OH radicals are produced and escape from the active site channel. This finding implies that the overexpression of the Cu,Zn-SOD gene (SODl), such as in Down's syndrome, will result in the formation of "free" -OH radicals in vivo and may be, in part, a cause of the syndrome. Transition metal ions are generally required for the generation of oxygen radicals. We have studied the effects of Mn(II) ions in physiological concentrations of bicarbonate and carbon dioxide. During the disproportionation of hydrogen peroxide catalyzed by Mn(II), superoxide and hydroxyl radicals are detected. Addition of an amino acid in the solution resulted in production of an amino acid-derived radical that replaced the superoxide radical. By employing various isotope-enriched amino acids we have identified this radical as a hydronitroxide -OOCC(R)HNHO-. The data are consistent with the formation of a transient "caged" -OH in the inner coordination sphere of Mn(II). Two reaction schemes are proposed to account for the experimental results.